Under basal conditions, the majority of glucose is metabolized through the glycolytic pathway, but 2-4 % is metabolized via the hexosamine biosynthetic pathway (HBP). The HBP modifies glucose to produce an O-linked N-acetylglucosamine (O-GlcNAc) moiety that can be added to serine/threonine residues of proteins in a highly dynamic and reversible reaction. Flux through the HBP is increased when glucose levels are in excess, which can lead to a pathological increase of O-GlcNAcylated proteins. Two enzymes, O-GlcNAc transferase (OGT) and O-GlcNAcase are responsible for adding and removing, respectively, the O-GlcNAc moiety to serine/threonine residues in proteins. Hippocampal neurons have the highest expression of O- GlcNAc transferase (OGT) and O-GlcNAcase in forebrain. Long-term changes in the efficacy of CA3-CA1 synapses underlie hippocampal dependent learning. The high expression of OGT and O-GlcNAcase in hippocampus suggests that normal synaptic function in this brain region is modulated by O-GlcNAc turnover of synaptic proteins. However little to nothing is known regarding how O-GlcNAcylation modulates synaptic function. Furthermore, the possibility exists that abnormal addition of O-GlcNAc on synaptic proteins could interfere with the ability of synapses to express long-term plasticity required for memory processing and could explain deficits in hippocampal synaptic function and learning known to occur in animal models of diabetes, where O-GlcNAcylation is pathologically elevated. No study to date has investigated the effects of O-GlcNAcylation on memory formation, either under physiological or pathological conditions. In recent studies, we find that OGT and O-GlcNAcase are tonically active and bidirectionally modulate the strength of basal synaptic transmission, suggesting the natural flux through the HBP sets the level of excitability in the circuit. Furthermore, we find that an increase in O-GlcNAcylation limits the ability of synapses to express normal LTP, with no effect on LTD. In this proposal we will investigate the cellular and molecular mechanisms mediating the synaptic depression induced by increased O-GlcNAcylation and determine whether chronic increases in O-GlcNAcylation causes synaptic dysfunction and learning deficits. Thus, the successful demonstration of a physiological role of O-GlcNAcylation in modulating synaptic transmission and plasticity could be the next major discovery in the field of learning and memory. The results obtained will launch a new area of investigation aimed at understanding how fluctuations in glucose metabolism by the HBP can directly affect synaptic function in physiological and pathological conditions. PUBLIC HEALTH RELEVANCE: The proposed studies are aimed at understanding how the highly dynamic posttranslational modification, O-GlcNAcylation, modulates synaptic function and the ability of hippocampal synapses to undergo long-term changes in synaptic efficacy the underlie learning and memory. This is a highly novel area of investigation in neurons that is likely to be as important in synapse function as phosphorylation. The results of these studies could provide a mechanistic understanding of deficits in synaptic plasticity and learning, consequences of diabetes and Alzheimer's disease, where O-GlcNAcylation is pathologically increased and decreased, respectively.